All publications mentioned throughout this application are fully incorporated herein by reference, including all references cited therein.
Cancer of all forms is one of the major causes of morbidity throughout the world. Research in the area of cancer chemotherapy has produced a variety of antitumor agents which have differing degrees of efficacy. A variety of cancer therapeutic agents are known, for example, alkylating agents, antimetabolites, alkaloids and carcinostatic antibiotics. Standard clinically used agents include adriamycin, actinomycin D, methotrexate, 5-fluorouracil, cis platinum, vincristine and vinblastine. However, the presently available antitumor agents are known to have various disadvantages such as toxicity to healthy cells and resistance of certain tumor types.
Hepatocellular carcinoma (HCC) is one of the major malignant diseases in the world today, the greatest incidence being in Japan, China, other parts of the Asia and sub-Saharan Africa. Recent evidence indicates that the incidence of hepatocellular carcinoma in Europe and North America is increasing. The disease is estimated to be responsible for, or involved in, up to approximately 1,250,000 deaths a year, and as such it is numerically one of the world's major malignant diseases.
The prognosis of HCC is poor, with the world-wide frequency rate almost equalling the mortality rate. After diagnosis, the median survival time is less than four months. Long-term survival, defined as survival longer than one year after the diagnosis, is seen only occasionally. Most HCC patients succumb to either the complications of liver failure with or without massive bleeding, or to the general effects of a large tumor burden, with cachexia, malnutrition, infection and sepsis. Though distant metastases occur (up to 90% of patients have metastatic tumors at the time of death), hepatic disease most often limits survival.
Current therapies available to the clinician are on the whole ineffective as a cure for this disease. For patients with advanced HCC who are not candidates for surgical resection, liver transplantation, localized tumor ablation or systemic chemotherapy remains the mainstay of therapy. Unfortunately, HCC is a relatively chemotherapy-resistant proliferative disorder; therefore, outcomes using this mode of treatment are unsatisfactory. Resistance to chemotherapy may be caused by the universal expression of the multidrug resistance gene protein on the surface of the malignant cells, leading to active efflux of chemotherapeutic agents. Chemotherapy is usually not well tolerated and seems to be less efficacious in patients with HCC with underlying hepatic dysfunction. Younger patients with well-compensated cirrhosis due to chronic hepatitis B or C infections have better outcome with chemotherapy than older patients with alcoholic cirrhosis and other comorbid diseases.
The most active single agent drugs tested have been doxorubicin, cisplatin, and fluorouracil. Response rates are about 10%, and treatment shows no clear impact on overall survival. More recently, gemcitabine and capecitabine have been evaluated in clinical trials, but response rates have been low and short term.
A variety of combination chemotherapy regimens have also been studied. Recently, cisplatin-based combination regimens have shown improved response rates around 20%, but to date, no survival advantage as compared to supportive care alone has been shown. No difference seems to exist in response rates between 2 or 3-drug regimens. Moreover, some of these combination regimens cause considerable toxicity.
Chemoimmunotherapy uses a combination of chemotherapy and immunomodulatory agents, such as interferon-alpha, to try to achieve better tumor response rates. Antiangiogenesis agents (i.e., bevacizumab), which work by disrupting the formation of blood vessels that feed tumors, are a new class of drugs that may prove to be of benefit in the treatment of HCC. The highly vascular nature of HCC tumors makes therapy with an antiangiogenesis agent a promising and exciting new option. Further evaluation of these drugs in the setting of clinical trials is needed to determine their efficacy.
Thus, there is a clear need for novel therapeutic approaches for specifically affecting cancerous cells, and especially, HCC. The present invention showed for the first time that ornithine aminotransferase is overexpressed in malignant proliferative tissue. The inventors further showed that inhibition of the enzyme catalytic activity using two different inhibitors, markedly decreased tumor growth, and therefore may be used as a specific target in the treatment of proliferative disorders, and particularly of HCC.
Ornithine aminotransferase (OAT) is a mitochondrial matrix enzyme that catalyzes a reversible reaction of interconversion between ornithine and alpha ketoglutarate to delta-1-pyrroline-5-carboxylate and glutamate. The enzyme is expressed in many tissues, including liver, kidney, small intestine, brain and eye. The enzymes from liver and kidney differ significantly in their regulation, and were believed to be two distinct enzymes. However, DNA sequencing proved that the two enzymes are encoded by a single gene.
As indicated above, glutamate is the product of the reaction catalyzed by OAT. This product can be used as a substrate by glutamine synthetase to synthesize glutamine, which is critical for the growth of proliferative cells, supporting protein and nucleotide synthesis and providing a major source of energy. Therefore an increased activity of OAT could make a tumor cell independent of any glutamine supply and confer a growth advantage to the cell. Thus, without being bound by any theory, it may be hypothesized that reducing the level of tissue glutamine concentrations by inactivation of OAT may lead to inhibition in cell proliferation and tumor growth.
Furthermore, it should be noted that ornithine is a substrate in the urea cycle. The urea cycle is effective in incorporating ammonium ions into urea in order to be eliminated from the body. The present invention is based on inhibition or inactivation of ornithine aminotransferase (OAT). Therefore, alternatively or additionally and without being bound by any theory, it may be postulated that by enhancing the level of tissue ornithine concentrations due to inactivation of OAT over an extended period of time, urea formation in the liver and presumably in some other tissues would be a consequence thereof, thereby lowering blood and cerebrospinal fluid ammonia concentrations. These compounds were implicated in numerous well known human illnesses associated with elevated blood and cerebrospinal fluid ammonia concentrations, among which, for example, are liver cirrhosis, fulminant hepatic failure and urinary tract/bladder infections. These, and particularly cirrhosis, may therefore lead to HCC.
Thus, the present invention comprises the new use of OAT inhibitors, for the treatment of proliferative disorders.
It is thus one object of the invention to provide a method for the treatment of proliferative disorders, and particularly HCC, which is based on inhibiting the expression or the activity of ornithine aminotransferase (OAT).
Another object of the invention is the use of ornithine aminotransferase inhibitors, and particularly of Gabaculine and Gabaculine analogue 8, for the preparation of pharmaceutical composition for the treatment of malignant proliferative disorders.
Another object of the invention is to provide for a diagnostic method for detection of proliferative disorders, particularly of HCC, by detecting increase in the expression of ornithine aminotransferase.
These and other objects of the invention will become apparent as the description proceeds.